Intradermal gene vaccination of mice with naked plasmid DNA has been previously shown by our laboratory to induce strong, specific immune responses characterized by the generation of MHC class I restricted CTL secretion of antigen specific IgG but not IgE, and a T helper response polarized toward the Th1 phenotype. Furthermore, gene vaccination can both prevent a subsequent IgE response to protein vaccination and down-regulate an ongoing IgE response. The overall objective of Project 3 is to identify the cellular components, cytokine mediators and their critical interactions elicited by intradermal plasmid DNA gene vaccination. Their roles in the initiation of this Th1 biased immune response, the prevention of IgE generation and the down-regulation of an ongoing IgE mediated response will be defined. In Aim 1, the relative roles of different lymphoid subsets: CD4+ T cells; CD8+ T cells; and gamma 8 T cells will be examined using either gene "knockout" mice, or in vivo subset depletion. In order to define the role of antigen presentation in the context of MHC class I and class II, we will make chimeric mice repopulated with bone arrow derived from ice lacking either beta2-microglobulin or MHC class II respectively. In Aim 2, the relative roles of the CD4+ , CD8+ and alphabeta and gamma 8 T cells subsets in the maintenance of the immune response will be assessed by adoptive transfer or primed populations to naive recipients. In Aim 3, the relative roles of the cytokines IFNgamma and IL-12 in the immune response to the gene vaccination will be determined using gene "knockout" mice. These experiments will initially be conducted using test antigen systems, but as allergen/vector combinations become available through Projects 1 and 2, the rules defined in the test antigen systems will be verified in Aim 4 for the allergen gene vaccines. Information generated by these experiments will be used to develop more refined vaccination strategies aimed at inducing a Th1 response and reducing an IgE response to allergens.